A. Field of the Invention
The invention relates, in an initial aspect, to the detection and treatment of an autoimmune disease, myasthenia gravis. Compositions of matter (peptides and combinations of peptides) and methods are disclosed which enable a high degree of predictive capability for detecting this neurological disease. The invention's therapies rely, in the first instance, on compositions and methods which are sensitive enough to allow detection of low levels of the autoantibodies associated with the symptoms of the disease, yet which virtually eliminate false readings. With such sensitive and accurate tests, a physician may confidently proceed to apply the treatments and methods using the therapeutic compositions and methods of the invention.
The invention relates, in another aspect, to procedures for synthesis of well-defined conjugates of peptides covalently bonded to tolerogenic polymers such as monomethoxypolyethylene glycol (mPEG) or polyvinyl alcohol (PVA). The first step in said synthesis involves succinylation of free-hydroxyl groups on the tolerogenic polymer by reaction with succinic anhydride. The polymer is then coupled to one or the other terminus, for instance via the carboxyl of a succinyl group to the .alpha.-NH.sub.2, of a peptide. This is achieved while maintaining intact all the side-chain protecting groups on the peptide. The MPEG or PVA-peptide conjugates are cleaved from a synthetic resin and purified. This method results in the preparation of conjugates in which one molecule of tolerogenic polymer is specifically coupled to one or the other or both of the termini of an otherwise unaltered peptide molecule.
In order to test the ability of such tolerogenic peptides to suppress antibody responses in an autoimmune disease, a synthetic peptide, .alpha.125-148, corresponding to a myasthenogenic region of Torpedo californica acetylcholine receptor (AChR) was conjugated to monomethoxypolyethylene glycol (MPEG). Injection of mice with the MPEG-(.alpha.125-148) conjugate and subsequent immunization with whole Torpedo AChR suppressed the development of experimental autoimmune myasthenia gravis (EAMG or MG) by electrophysiological criteria. In anti-AChR antisera from these animals, the antibody response against the unconjugated peptide .alpha.125-148 was decreased while the antibody responses against whole AChR and other epitopes were not altered. There were no detectable changes in T cell proliferation responses to peptide .alpha.125-148 or to whole AChR in these animals. Prior injections with a "nonsense" peptide mPEG conjugate had no effect on responses to the subsequent immunization with whole Torpedo AChR. The results indicate that the mPEG-(.alpha.125-148) conjugate has epitope-specific tolerogenicity for antibody responses in EAMG, and that the region .alpha.125-148 plays an important pathophysiological role in EAMG. These studies strongly indicate that other epitope-directed tolerogenic conjugates will be useful for future immunotherapies of human myasthenia gravis.
By way of further example, tolerogenic peptides are also disclosed for diseases as diverse as ragweed pollen allergy and Grave's disease. The strategy of specific suppression of the antibody response to a pre-determined epitope using a synthetic mPEG-peptide conjugate will be useful in manipulation and suppression of unwanted immune responses such as autoimmunity and allergy.
B. Specific Background